This patent application claims a benefit of priority from Korean Patent Application No. 2000/20137 filed Apr. 17, 2000 and Korean Patent Application No. 2000/31926 filed Jun. 10, 2000 through PCT Application Serial No. PCT/KRO1/00613 filed Apr. 13, 2001, the contents of each of which are incorporated herein by reference.
The present invention relates to novel 6-methylnicotinamide derivatives and pharmaceutical compositions containing said derivatives. More specifically, the present invention relates to novel 6-methylnicotinamide derivatives and their pharmaceutically acceptable salts represented below in formula 1, which have an excellent inhibitory effect on proliferation of hepatitis B virus(HBV), hepatitis C virus(HCV) and human immunodeficiency virus(HIV). The present invention also relates to the process for preparing compounds of formula 1 and the pharmaceutical compositions containing said derivatives as effective ingredients against viruses. 
wherein,
R1 is hydroxy group; straight or branched C1xcx9cC5 alkyl group; C3xcx9cC6 cycloalkyl group which is unsubstituted or substituted with hydroxy group, C2xcx9cC6 dialkylamino group; saturated or unsaturated 5 or 6 membered heterocyclic compounds containing 1 to 2 heteroatoms selected from N, O and S which may be unsubstituted or substituted with C1xcx9cC3 alkyl group; or 
xe2x80x83m is an integer of 0 or 1,
R2 is H or C1xcx9cC4 alkyl group;
or R1 and R2 are joined to form a 5- or 6-membered heterocyclic ring containing 1 to 2 heteroatoms selected from N, O and S,
n is an integer from 0 to 4,
R3 is 5-indazole or 6-indazole
Hepatitis B virus (HBV; referred as xe2x80x9cHBVxe2x80x9d hereinafter) causes acute or chronic hepatitis, which may progress to liver cirrhosis and liver cancer. It is estimated that three hundred million people are infected with HBV in the world (Tiollais and Buendia, Sci. Am., 264, 48, 1991). There has been much research about the molecular biological characteristics of HBV and their relationship to liver diseases in order to find ways to prevent and treat hepatitis B. Various vaccines and diagnostic drugs have been developed and much effort is being channeled into research to find treatment for hepatitis B.
HBV genome consists of genes for polymerase (P), surface protein (pre-S1, pre-S2 and S), core protein (pre-C and C), and X protein. Of these proteins expressed from HBV genes, polymerase, surface protein, and core protein are structural proteins and X protein has a regulatory function.
The gene for HBV polymerase comprises 80% of the whole virus genome and produces a protein of 94 kD size with 845 amino acids, which has several functions in the replication of virus genome. This polypeptide includes sequences responsible for activities of protein primer, RNA dependent DNA polymerase, DNA dependent DNA polymerase, and RNase H. Kaplan and his coworkers first discovered reverse transcriptase activities of polymerase, which led to much research in replicating mechanism of HBV.
HBV enters liver when antigenic protein on virion surface is recognized by hepatic cell-specific receptor. Inside the liver cell, DNAs are synthesized by HBV polymerase action, attached to short chain to form complete double helix for HBV genome. Completed double helical DNA genome of HBV produces pre-genomic mRNA and mRNAs of core protein, surface protein, and regulatory protein by the action of RNA polymerase. Using these mRNAs, virus proteins are synthesized. Polymerase has an important function in the production of virus genome, forming a structure called replicasome with core protein and pre-genomic mRNA. This process is called encapsidation. Polymerase has repeated units of glutamic acid at the 3xe2x80x2-end with high affinity for nucleic acids, which is responsible for facile encapsidation. When replicasome is formed, (xe2x88x92) DNA strand is synthesized by reverse transcribing action of HBV polymerase and (+) DNA strand is made through the action of DNA dependent DNA polymerase, which in turn produces pre-genomic mRNAs. The whole process is repeated until the pool of more than 200 to 300 genomes is maintained (Tiollais and Buendia, Scientific American, 264: 48-54, 1991).
Although HBV and HIV are different viruses, the replication mechanisms during their proliferation have some common steps, namely, the reverse transcription of virus RNA to form DNA and the removal of RNA strand from subsequently formed RNA-DNA hybrid.
Recently, nucleoside compounds such as lamivudine and famvir have been reported to be useful inhibitors of HBV proliferation, although they have been originally developed as therapeutics for the treatment of acquired immune deficiency syndrome (AIDS; referred as xe2x80x9cAIDSxe2x80x9d hereinafter) and herpes zoster infection (Gerin, J. L, Hepatology, 14: 198-199, 1991; Lok, A. S. P., J. Viral Hepatitis, 1: 105-124, 1994; Dienstag, J. L. et al., New England Journal of Medicine, 333: 1657-1661, 1995). However, these nucleoside compounds are considered a poor choice for treatment of hepatitis B because of their high cost and side effects such as toxicity, development of resistant virus and recurrence of the disease after stopping treatment. Effort to find therapeutics for hepatitis B among non-nucleoside compounds has been continued and antiviral effects against HBV have been reported for quinolone compounds (EPO 563732, EPO 563734), iridos compounds (KR 94-1886), and terephthalic amide derivatives (KR 96-72384, KR 97-36589, KR 99-5100). In spite of much effort, however, effective drugs for treating hepatitis B have not been developed yet and therapeutic method mainly depends on symptomatic treatment.
Hepatitis C virus (referred as xe2x80x9cHCVxe2x80x9d hereinafter) is a virus of the flaviviridae family which has a membrane. HCV genome is single stranded (+)-RNA of 9.5 kb in length and express polyprotein comprising of 3010 amino acids. The HCV polyprotein is cleaved co- and post translationally by cellular and viral protease to yield 3 structural proteins and 6 nonstructural proteins. 5xe2x80x2- and 3xe2x80x2-terminus of the HCV genome contain untranslated region (UTR), which highly conserved nucleotide sequence of all most genotype. Recently, it is known that 5xe2x80x2-UTR is a 330xcx9c341 nucleotide sequences and 3xe2x80x2-UTR includes 98 nucleotides at the back of poly A, termed to X region which might be played a role of RNA replication and post-translation of virus. Amino end part of HCV genome produces structural proteins Core, E1 and E2 and the other part comprise of non-structural protein. The core is the main structural component of the viral capsid and E1 and E2 comprises of a outer protein. These proteins are cleaved by signal peptidase in endoplasmic reticulum. Serin-type protease NS3 and cofactor NS4A are responsible for the cleavage of nonstructural protein. NS5B protein is a RNA-dependant RNA polymerase. This protein is the most importance enzyme involved in the regulation of HCV replication.
It is reported that an infection by HCV is generated from a blood transfusion and community-acquired infection. Approximately 70% of HCV infected individuals will develop chronic hepatitis, of which 20% will progress to severe chonic liver disease within 5 years. Such higher progression rate, rarely in RNA virus, shows that HCV is a major cause of generating liver cancer. Mechanism studies of the continuous infection of HCV have not been reported. HCV test is therefore carried out in all blood and the infection opportunity by the blood transfusion is remarkably decreased. But, HCV infection presents a major public health problem worldwide because the community-acquired HCV infection has not regulated yet.
From the view of retrospective studies, HCV infection uniformly distributes worldwide and 1.5-2% of the world""s population is infected. Compared to HBV, HCV infection is generally developed in chronic hepatitis and has a high probability of progression to liver cirrhosis and liver cancer. Hepatitis C virus that belongs to completely different family cannot be inhibited using B-type vaccine and treatment with xcex1-interferon. That treatment have not shown a remarkable antiviral effect owing to their variable genotype.
Since HCV was discovered in 1987, there has been attempted a lot of research, but remarkably effective drug has not yet developed. Interferon is the unique choice for the treatment so far, but it has confirmed that its medical care rate is less than 30%, HCV is recurred after cessation of its treatment and several interferon-resistant mutant virus generates. So far, there are not characteristic antiviral agents with proliferation inhibitory activity against HCV.
Meanwhile, AIDS is a disease inducing dramatic decrease in immune function in the body cells and causing various symptoms of infection rarely seen in normal human beings, which spread to the whole body. Human immunodeficiency virus (HIV; referred as xe2x80x9cHIVxe2x80x9d hereinafter) responsible for AIDS is known to mainly attack helper T cells, which is one of the T cells with regulatory function in the immune system. When helper T cells are infected with HIV virus and undergo necrosis, human immune system cannot function properly. Impairment in immune function subsequently results in fatal infection and development of malignant tumor. Since AIDS patient has been found in USA in 1981 for the first time, the number increased to more than 830,000 patients in 187 countries in 1993 (WHO 1993 report). WHO predicted that 30 to 40 million more people would be infected with HIV by the year 2000 and 10 to 20 million of them would develop the disease.
At the present time, drugs controlling proliferation of HIV have been most widely used for the treatment of AIDS. Of these, Zidovudine, which had been named Azidothymidine previously, is a drug developed in 1987. Didanosine was developed in 1991 as an alternative medicine for AIDS patients when Zidovudine was either ineffective or could not be used due to side effects. In addition, Zalcitabine was approved for concurrent use with Zidovudine in 1992. These drugs alleviate symptoms, slow down progression of the disease in the infected individuals to full-blown AIDS, and somewhat extend life span in the patients. These drugs, however, are not able to cure the patients completely and often develop problems such as resistance and side effects.
In light of these problems, we, inventors of the present invention, tried to develop therapeutics to treat hepatitis B with little chance of toxicity, side effects, and development of resistant viral strains. We found the compounds with excellent antiviral effect against HBV; synthesized novel 6-methylnicotinamide derivatives represented in formula 1 and completed the invention by showing their dramatic inhibitory effect on proliferation of HIV and HCV as well as of HBV.
It is an object of this invention to provide novel 6-methylnicotinamide derivatives, their pharmaceutically acceptable salts, and their preparation.
It is a further object of this invention to provide a pharmaceutical compositions containing said derivatives with cost effectiveness and little chance of side effects, as a therapeutic agent as well as a preventive agent for hepatitis B, hepatitis C and acquired immune deficiency syndrome (AIDS).
The present invention provides novel 6-methylnicotinamide derivatives represented by following formula 1 and their pharmaceutically acceptable salts. 
wherein,
R1 is hydroxy group; straight or branched C1xcx9cC5 alkyl group; C3xcx9cC6 cycloalkyl group which is unsubstituted or substituted with hydroxy group, C2xcx9cC6 dialkylamino group; saturated or unsaturated 5 or 6 membered heterocyclic compounds containing 1 to 2 heteroatoms selected from N, O and S which may be unsubstituted or substituted with C1xcx9cC3 alkyl group; or 
R2 is H or C1xcx9cC4 alkyl group;
or R1 and R2 are joined to form a 5- and 6-membered heterocyclic ring containing 1 to 2 heteroatoms selected from N, O and S,
n is an integer from 0 to 4,
R3 is 5-indazole or 6-indazole
More preferably, wherein,
R1 is hydroxyl, methyl, isopropyl, t-butyl, cyclo propyl, 4-hydroxyl-1-cyclohexyl, dimethyl amino, 4-morphorinyl, 1-piperidinyl, 4-methyl-1-piperazinyl, 2-pyridyl, 3-pyridyl, 1-imidazolyl, 2-pyridylamino or 3-pyridylcarbonyl amino,
R2 is H,
n is an integer from 0 to 3.
More preferable compounds in accordance with the present invention are as follows;
1) 2-(1H-5-indazolylamino)-N-isopropyl-6-methylnicotinamide, (compound of example 1),
2) 2-(1H-5-indazolylamino)-6-methyl-N-(4-morpholinyl)nicotinamide, (compound of example 2),
3) N-(trans-4-hydroxy-1-cyclohexyl)-2-(1H-5-indazolylamono)-6-methylnicotinamide, (compound of example 3),
4) 2-(1H-5-indazolylamino)-6-methyl-N-(3-pyridyl)nicotinamide, (compound of example 4),
5) 2-(1H-5-indazolylamino)-6-methyl-N-(3-pyridylmethyl)nicotinamide, (compound of example 5),
6) 2-(1H-5-indazolylamino)-6-methyl-N-(2-pyridylethyl)nicotinamide, (compound of example 6),
7) 2-(1H-6-indazolylamino)-6-methyl-N-methylnicotinamide, (compound of example 7),
8) 2-(1H-6-indazolylamino)-N-isopropyl-6-methylnicotinamide, (compound of example 8),
9) N-(t-butyl)-2-(1H-6-indazolylamino)-6-methylnicotinamide, (compound of example 9),
10) N-(2-hydroxyethyl)-2-(1H-6-indazolylamino)-6-methylnicotinamide, (compound of example 10),
11) N-cyclopropyl-2-(1H-6-indazolylamino)-6-methylnicotinamide, (compound of example 11),
12) N-[2-(dimethylamino)ethyl]-2-(1H-6-indazolylamino)-6-methylnicotinamide, (compound of example 12),
13) N-[2-(diethylamino)ethyl]-2-(1H-6-indazolylamino)-6-methylnicotinamide, (compound of example 13),
14) 2-(1H-6-indazolylamino)-6-methyl-N-[2-(4-morpholinyl)ethyl]nicotinamide, (compound of example 14),
15) 2-(1H-6-indazolylamino)-6-methyl-N-[2-(1-piperidinyl)ethyl]nicotinamide, (compound of example 15),
16) 2-(1H-6-indazolylamino)-6-methyl-N-(4-morpholinyl)nicotinamide, (compound of example 16),
17) 2-(1H-6-indazolylamino)-6-methyl-N-(4-methyl-1-piperazinyl)nicotinamide, (compound of example 17),
18) 2-(1H-6-indazolylamino)-6-methyl-N-(3-pyridyl)nicotinamide, (compound of example 18),
19) 2-(1H-6-indazolylamino)-6-methyl-N-(3-pyridylmethyl)nicotinamide, (compound of example 19),
20) N-[3-(1H-1-imidazolyl)propyl]-2-(1H-6-indazolylamino)-6-methylnicotinamide, (compound of example 20),
21) 2-(1H-6-indazolylamino)-6-methyl-N-(2-pyridyl)-3-pyridinecarbohydrazide, (compound of example 21),
22) 2-(1H-6-indazolylamino)-6-methyl-N-(3-pyridylcarbonyl)-3-pyridinecarbohydrazide, (compound of example 22),
23) 2-(1H-6-indazolylamino)-6-methyl-3-(4-morpholinylcarbonyl)pyridine, (compound of example 23).
The compounds according to the present invention have an inhibitory effect on proliferation of HIV as well as HBV, HCV since containing 5-indazolylamino or 6-indazolylamino as a substituent of 6-methylnicotinamide derivatives.
The compounds represented by formula 1 of the present invention may be utilized in the form of salts and the acid addition salts prepared by adding pharmaceutically acceptable free acids are useful. Compounds of formula 1 may be changed to the corresponding acid addition salts according to the general practices in this field. Both inorganic and organic acids may be used as free acids in this case. Among inorganic acids, hydrochloric acid, hydrobromic acid, sulfuric acid, or phosphoric acid may be used. Among organic acids, citric acid, acetic acid, lactic acid, tartaric acid, maleic acid, fumaric acid, formic acid, propionic acid, oxalic acid, trifluoroacetic acid, benzoic acid, gluconic acid, methanesulfonic acid, glycolic acid, succinic acid, 4-toluenesulfonic acid, galacturonic acid, embonic acid, glutamic acid or aspartic acid may be used.
The present invention also provides a process for preparing 6-methylnicotinamide derivatives of formula 1, represented by scheme 1 as follows: 
wherein, R1, R2, R3 and n are as defined in formula 1.
The process for preparing in the present invention comprises the following steps of:
1) preparing of 6-methylnicotinic acid derivatives (4) substituted with aminoindazole by heating and reacting 2-chloro-6-methylnicotinic acid (2) with 5-aminoindazole or 6-aminoindazole (3) in the presence of a base (step 1); and,
2) preparing of 6-methylnicotinamide derivatives of formula 1 by activating the 6-methylnicotinic derivatives (4) prepared in step 1, and then reacting the compounds of formula 4 with amine compound (5) (step 2).
A detail description will be stepwise given of the method for preparing 6-methylnicotinamide derivatives of the present invention.
Chemical reagents used as starting and reaction materials in the step 1 and 2 of the scheme 1, namely, 2-chloro-6-methylnicotinic acid (2), 5-aminoindazole (3), 6-aminoindazole (3) and amine compound (5), are commercially available and may be purchased.
The amine compound (5) in the step 2 is also used to introduce a constituent (R-NR2-(CH2)n-R1) into the desired compound of formula 1 and an appropriate amine compound should be selected depending on the substituent desired, which can be easily done by one with general knowledge in the technical field.
In the step 1, it is preferably used tertiary organic base having weak basicity such as pyridine, picoline, lutidine, N,N-dimethylaniline, 4-dimethylaminopyridine, as a base.
In the step 2, compounds of the formula 1 is prepared by reacting with amine compound (5) after activating 6-methylnicotinic acid derivatives (4) obtained by the step 1.
In activating process, 6-methylnicotinic acid derivatives are activated to highly reactive acid chloride compounds by adding a bit excess thionyl chloride to 6-methylnicotinic acid derivatives (4), subsequently heating, or, by Vilsmeier reagent prepared by heating thionyl chloride and N,N-dimethylformamide.
For a solvent, a protonic solvent is preferable, for example, a single or a mixture of solvents selected from chloroform, methylene chloride, acetonitrile, tetrahydrofuran and ether.
The reaction temperature is preferably 0xcx9c20xc2x0 C.
Furthermore, the present invention provides the pharmaceutical compositions of therapeutics comprising 6-methylnicotinamide derivatives or their pharmaceutically acceptable salts of formula 1 as effective ingredients to prevent and treat hepatitis B.
The present invention also provides the pharmaceutical compositions of therapeutics, comprising 6-methylnicotinamide derivatives or their pharmaceutically acceptable salts of formula 1 as effective ingredients to prevent and treat hepatitis C.
An inhibitor of RNA-dependant RNA-polymerase in the HCV, can essentially inhibit on HCV proliferation, and the compounds of the present invention is therefore expected to be useful for therapeutic agents on Hepatitis C with low toxicity since the RNA-dependant RNA-polymerase is not detected yet in human body cell.
6-methylnicotinamide derivatives of formula 1 in present invention have an inhibitory effect on proliferation of both HIV and HBV because they interfere with removal of RNA strand from RNA-DNA hybrid formed during the reverse transcription of viral RNA to DNA, which is a common step in the replication mechanism of the two viruses.
Therefore, the present invention also provides the pharmaceutical compositions of therapeutics for preventing and treating AIDS, which comprise 6-methylnicotinamide derivatives or their pharmaceutically acceptable salts of formula 1 as effective ingredients.
Compounds of formula 1 may be taken orally as well as through other routes in clinical uses; for example, it may be administered intravenously, subcutaneously, intraperitoneally, locally and in the form of general drugs.
For clinical use of drugs with the pharmaceutical compositions of the present invention, compounds of formula 1 may be mixed with pharmaceutically acceptable excipients and made into various pharmaceutically acceptable forms; for example, to tablets, capsules, trochese, solutions, suspensions for oral administration; infection solutions, suspensions, and dried powder to be mixed with distilled water for the formulation of instant injection solution.
Effective dosage for compound of formula 1 is generally 10xcx9c500 mg/kg, preferably 50xcx9c300 mg/kg for adults, which may be divided into several doses, preferably into 1xcx9c6 doses per day if deemed appropriate by a doctor or a pharmacist.
Hereinafter the present invention describes in more detail. However, it will be appreciated that those skilled in the art, on consideration of this disclosure, may make modifications and improvements within the spirit and scope of the present invention.